The field of selective delivery of biologically active molecules to specific cellular targets is rapidly advancing. Among the various systems that have been studied for their ability to deliver bioactive molecules to cells, lipoproteins, which form naturally occurring biological emulsions, are considered attractive as delivery vehicles for a number of reasons: 1) as endogenous carriers of cholesterol and other lipids in the blood circulation, they are not immunogenic and escape recognition by the reticuloendothelial system; 2) physicochemical characterization of drug-loaded lipoproteins indicates that particles with the same physicochemical properties as the native lipoproteins can be obtained; 3) lipoproteins are removed from the circulation by specific receptors that recognize their apoproteins or, they may be directed to nonlipoprotein receptors by chemical modification of their apoproteins (see, for example, Bijsterbosch, M. K. and van Berkel, T. J. C. (1990) Adv. Drug Delivery Revs, 5:231-251); 4) lipoproteins are physically stable due to their compact, neutral, apolar core; 5) the apolar core of lipoproteins provides an ideal domain for lipophilic molecules since molecules that are transported in the core of the lipoprotein are protected from the environment during transportation and the environment is protected from the molecule; and 6) lipoproteins can be synthesized from commercially available lipids and isolated apoproteins.
Examples of the use of lipoproteins as delivery vehicles for bioactive molecules include U.S. Pat. Nos. 4,868,158 and 5,324,821 which refer to the preparation of lipoproteins modified by incorporation of a lipophilic active substance into their apolar core. However, since many of the biologically active molecules used for treatment and/or prevention of diseases are too hydrophilic for incorporation into the apolar core of lipoproteins, investigators have attempted to find methods which would permit the incorporation of hydrophilic molecules into lipoproteins.
One approach that has been utilized is to couple hydrophilic molecules to hydrophobic anchors in order to render the hydrophilic molecule more lipophilic. For example, van Berkel has reported the synthesis of a dioleoyl derivative of the anti-viral nucleoside analogue iododeoxyuridine and its incorporation into either high density lipoproteins (HDL) (Bijsterbosch, M. K. et al. (1994) Biochemistry, 33:14073-14080) or chylomicrons (Rensen, P. C. N. et al. (1995) Nature Medicine, 1:221-225). Unfortunately, the use of conjugation to facilitate incorporation of hydrophilic molecules into lipoproteins has a number of potential drawbacks: 1) since the conjugated molecules are typically incorporated into a lipid or a protein residue on the surface of the lipoproteins, the conjugated molecule may interfere with the interaction of the apoprotein of the lipoprotein and its receptor; 2) such surface-modified particles may show a greater tendency to aggregate due to a potential loss of surface charge; 3) attachment of bioactive molecules to the surface of lipoproteins exposes the molecules to the environment and vice versa; 4) “derivatization” of hydrophilic molecules by conjugation to hydrophobic anchors may affect the biological activity of the hydrophilic molecule; and 5) such a “conjugation” approach is impractical for rendering larger molecular weight hydrophilic molecules hydrophobic enough to be incorporated into the apolar core of the lipoproteins.